Mutation Spectrum Induced by 8-Bromoguanine, a Base Damaged by Reactive Brominating Species, in Human Cells

Figure 3

Excision of thymine mispaired with 8-bromoguanine (8BrG) by SMUG1 and TDG proteins. (a) Time-course assay for the cleavage of 30-mer double-stranded oligonucleotides containing a T:8BrG mispair by SMUG1 and TDG proteins. Each protein or no DNA glycosylase proteins (−) were incubated at 37°C for 0–120 min with a T:8BrG-containing oligonucleotide. The amount of cleavage products as a proportion of the total oligonucleotides was calculated as the % incision. The % incision values were shown as the means ± standard deviations of data from three independent experiments. The lower panels show representative results of the DNA cleavage activity assays. (b) Protein concentration dependency of cleavage of double-stranded oligonucleotide containing a T:8BrG mispair by SMUG1 and TDG proteins. Each protein was incubated at 37°C for 60 min with a T:8BrG-containing oligonucleotide. The amount of cleavage products as a proportion of the total oligonucleotides was calculated as the % incision. Data are shown as the means ± standard deviations. The right panel shows a representative result. “I” and “C” indicate intact oligonucleotides and cleaved oligonucleotides, respectively (a and b).

(a)

(b)